蜕膜NKT细胞过度表达穿孔素与流产发病的关系

采用腹腔注射α-半乳糖基神经酰胺(α-galactosylceramide,α-GC)的方法诱发C57BL/6小鼠流产,并检测注射后小鼠蜕膜NKT细胞表达穿孔素的情况,进而分析α-GC导致流产的发病机制。研究发现,在孕6.5 d腹腔注射α-GC(100μg/kg体重)可使小鼠孕10.5 d胚胎吸收率达到35.6%(37/104),显著高于对照组(7.4%;10/135;P<0.01)。此时采用流式细胞术检测蜕膜CD3+CD49b+NK1.1+细胞表达穿孔素的水平,发现α-GC刺激后穿孔素平均检出率显著高于对照组(15.5%和4.6%;P<0.01)。预先注射穿孔素的选择性抑制剂乙二醇四乙酸(ethylene glycol tetraacetic acid,EGTA)抑制穿孔素的作用,可显著降低α-GC所引起的小鼠胚胎吸收率增高。EGTA处理组和溶剂对照组α-GC诱导后小鼠胚胎吸收率分别为13.4%(16/119)和40.8%(42/103;P<0.01)。这些结果提示,α-GC可上调NKT细胞穿孔素的表达,而过多的穿孔素可能是α-GC导致小鼠胚胎吸收率增高的分子基础。

Relationship of the overexpression of perforin by decidual NKT cells and the pregnancy loss

To explore the mechanism by which α-galactosylceramide(α-GC) induces pregnancy loss.The C57BL/6×BALB/c mouse model was injected with α-GC intraperitoneally on 6.5 days post-gestation(6.5 d) into pregnant mice to induce embryo resorption.The status of perforin expression in NKT cells was determined by flow cytometry assay using CD3+CD49b+ cells purified by magnetic affinity cell sorting(MACS).It was found that the resorption rate of embryos on 10.5 d was significantly increased after α-GC injection at a dosage of 100 μg/kg body weight,it was 35.6%(37 of 104) in the α-GC-treated group in comparison with 7.4%(10 of 135) in the PBS control group(P<0.01).The proportion of perforin-producing cells in the MACS-purified CD3+CD49b+NK1.1+ cell population was markedly higher in response to α-GC induction(15.5% vs 4.6%,P<0.01).In addition,previous injection of ethylene glycol tetraacetic acid(EGTA),a selective perforin inhibitor,dramatically diminished the increase of embryo loss induced by α-GC.The resorption rate of embryos was significantly lower in EGTA-inhibited group(13.4%;16 of 119) than no-inhibition group upon α-GC-induction(40.8%;42 of 103;P<0.01).These results suggest that α-GC may up-regulate perforin production in decidual NKT cells and increased murine embryo loss may be attributed to overexpression of perforin.